Malaria is caused by parasites of the genus Plasmodium and causes an estimated 300-500 million clinical cases and 1-3 million deaths annually (Snow et al, 2005). In addition to morbidity and mortality the economic burden due to malaria is immense, with loss of up to 1-2% of GDP per year estimated for some countries where malaria is endemic.
An essential step in the life cycle of malaria parasites is the invasion of host erythrocytes by merozoites. The invasion process is characterized by a multitude of specific, but relatively poorly understood, interactions between protein ligands expressed by the merozoite and receptors on the erythrocyte surface (Cowman A F and Crabb B S, 2006). Several molecules implicated in the invasion process have been identified in the apical organelles (rhoptry, micronemes, and dense granules) of the merozoite. At least two gene families the Reticulocyte Binding Protein homologues (RH) and the family of erythrocyte binding proteins/ligands (EBL) have been shown to mediate specific interactions with host cell receptors thereby defining host cell specificity and are thought to play an important role in parasite virulence and possibly immune evasion (Gaur et al., 2004; Iyer et al., 2007).
In the human parasite Plasmodium falciparum two gene families termed Erythrocyte Binding Like Proteins (or EBL) and the Reticulocyte Binding Protein Homologues (RBPH) have been shown to play a crucial role in the selection of suitable host cells. Both EBL and RBPH are thought to directly interact with specific receptors on the red blood cell surface. In the case of EBL the region within the protein that directly mediates binding has been identified. This region called Duffy Binding Like Domain (DBL) is characterized by a number of conserved cysteine residues and is conserved in all members of this gene family. In contrast no binding region of any RBPH member has so far been identified. Considering the large size of these proteins (up to 300 kDa) it is crucial to dissect the protein into smaller functional domains.
Numerous studies have indicated that malarial merozoites can invade erythrocytes through several invasion pathways. This ability is dependent on the repertoire of parasite ligands expressed at the surface of the parasite and variations of receptors at the erythrocyte surface. The various alternative invasion pathways are classified according to the nature of the erythrocyte receptors involved in invasion, which in turn are operationally defined by the enzymatic treatments upon erythrocytes which disrupt binding. P. falciparum EBA-175, one of the EBL members, is the best characterized receptor and recognizes sialic acid components on Glycophorin A (Sim B K L et al, 1994). Other EBLs have been shown to interact with a Glycophorin B and C as well as the Duffy blood group antigen.
In P. falciparum five RH members PfRH1, PfRH2a & 2b, PfRH3, and PfRH4 have been identified (Cowman A F and Crabb B S, 2006). Recognition of erythrocytes by PfRH1 is sialic acid dependent and trypsin resistant (Rayner J C et al, 2001), whereas that of PfRH2b is sialic acid independent and trypsin resistant (Duraisingh et al., 2003), and that of PfRH4 is sialic acid independent and trypsin resistant (Stubbs et al, 2005). While all these studies indicate that RH recognizes a specific receptor on the erythrocyte surface, only in the case of PfRH1 has direct binding to red blood cells been demonstrated.
How binding of EBL or RH to specific erythrocyte receptors ultimately leads to merozoite invasion is an important question that requires the parasite ligand to be dissected into functional domains.